Models for the ultimate development of effective vaccines and immunotherapies that would limit HIV replication can be drawn from naturally occurring examples of immune system-mediated control. Identifying the components, targets, and magnitude of an effective immune response to HIV are important steps toward developing effective vaccines and immunotherapies. Although patients with normal CD4+ T cell counts and low levels of plasma virus are a heterogeneous group, a small subgroup of patients with truly non-progressive HIV infection and restriction of virus replication likely holds important clues to the basis of an effective immune response to HIV. A small subpopulation of HIV-infected individuals (fewer than 0.8%) shows no signs of disease progression over a 10-year period. We have assembled a stringently defined cohort of such patients, termed long-term nonprogressors (LTNPs), or elite controllers. Many of these patients have been infected for 20 years, yet even without receiving antiretroviral therapy, they have experienced no CD4+ T-cell decline and have maintained plasma viral RNA levels below 50 copies per milliliter. We are using cells from these patients to systematically dissect the mechanisms of immune-mediated restriction of virus replication. The HIV-specific T-cell responses of these patients have been studied in extreme detail. Through this project, considerable progress has been made in understanding how the immune system controls HIV. Our prior work indicated that there is a dramatic association between immunologic control and the HLA B*5701 allele, and that the immune response is highly focused on peptides restricted by this allele. This result established both host genetic and functional links between immunologic control and the CD8+ T-cell responses of these patients. The finding of high frequencies of CD8+ T cells specific for the patients virus in both LTNPs and progressors strongly suggested that differences between responses of these patient groups were qualitative rather than quantitative in nature. One important qualitative difference in the HIV-specific immune response that distinguishes LTNPs from progressors is the maintenance of HIV-specific CD8+ T cells with a high proliferative capacity. This proliferation parallels perforin expression required for effective killing of HIV-infected CD4+ T cells. To better understand the mechanisms of immunologic control, we devised a method to measure HIV-infected cell elimination on a per-cell basis. Measured on a per-cell basis, HIV-specific CD8+ T cells of LTNPs efficiently eliminated primary autologous HIV-infected CD4+ T cells. This effective killing was clearly distinguishable from the responses of progressors over a very broad range of effectors to HIV-infected targets. Progressor cells did not mediate effective killing even at high effector-to-target ratios. Defective cytotoxicity of progressor effectors could be restored in vitro. These results establish an effector function and a mechanism that clearly segregate with immunologic control of HIV. One of the original goals of our work in LTNP was to provide insights regarding important elements of the cellular immune response that should be induced in vaccination. Although cytotoxic capacity clearly distinguishes those with immunologic control in the setting of chronic infection, it may not necessarily be the operative mechanism in control induced by an HIV vaccine. Previously, direct measurements of recall cytotoxic capacity assessed by granzyme B target cell activity and infected CD4+ T-cell elimination had not been applied to recipients of HIV vaccines. We recently examined the HIV-specific CD8+ T-cell cytotoxic capacity of HIV-1-uninfected recipients of the Merck Ad5/HIV trivalent vaccine. A very high profile efficacy trial employing this vaccine was the Step study, a phase IIB test-of-concept trial involving 3,000 HIV-negative individuals at high risk of HIV infection. We observed that the ability of vaccine-induced HIV-specific CD8+ T cells to kill primary autologous HIV-infected targets was relatively low and not comparable to responses we have previously associated with immunologic control. These findings suggest that the relatively poor ability of this vaccine to reduce viral loads upon natural infection is potentially attributable to the relatively poor induction of cytotoxic capacity. The next step in this project is to determine whether cytotoxic capacity is predictive of the ability to control HIV in the few vaccinees that exhibited control of the virus upon natural infection. We have recently extended our observations in humans to Simian Immunodeficiency Virus(SIV)-infected Rhesus macaque model. This is the most widely used primate model of HIV infection and offers the possibility of studying cytotoxic capacity in a model where there are excellent examples of immunologic control, either spontaneous or following vaccination. In addition this model offers the possibility of studying the causal relationships between cytotoxic capacity and immunologic control in a manner that cannot be pursued in humans. A subset of SIV-infected rhesus macaques behave as LTNP manifesting similar features of effective immune system-mediated control of lentiviral infection. MHC class I alleles are associated with control of SIV infection, particularly Mamu B*08 and B*17. The CD8+ T cells of LTNP carrying these alleles preferentially recognize Mamu B08 and B17-bound SIV epitopes. Furthermore, the 2-4 log increase in SIV plasma viremia seen after in vivo CD8+ T cell depletion in both LTNP and progressors in vivo provides further support that SIV is controlled by the CD8+ T cell response in these animals. Although the study of non-human primates has resulted in important advances for understanding HIV-specific immunity, a clear correlate of immune control over SIV replication had not been found. In 2013, CD8+ T-cell cytotoxic capacity was examined to determine whether this function is a correlate of immune control in the rhesus macaque SIV infection model as has been suggested in chronic HIV infection. SIVmac251-infected CD4+ T-cell clone targets were co-incubated with autologous macaque effector cells to measure infected CD4+ T-cell elimination (ICE). Twenty-three SIV-infected rhesus macaques with varying plasma viral RNA levels were evaluated in a blinded fashion. LTNP had greater median ICE than progressors (67.3% 22.0-91.7% vs. 23.7% 0.0-58.0%, p=0.002). In addition, significant correlations between ICE and viral load (r= -0.57, p=0.01), and between granzyme B delivery and ICE (r=0.89, p<0.001) were observed. Furthermore, the CD8+ T cells of LTNP exhibited greater per-cell cytotoxic capacity than those of progressors (p=0.004). These findings suggest that greater lytic granule loading of virus-specific CD8+ T cells and delivery of active granzyme B to SIV-infected targets are associated with superior control of SIV infection in rhesus macaques, consistent with observations of HIV infection in humans. Therefore, such measurements appear to represent a correlate of control of viral replication in chronic SIV infection and their role as predictors of immunologic control in the vaccine setting should be evaluated. Taken together our work suggests that cytotoxic capacity is a clear correlate of immunologic control of HIV in chronic infection and may also be an important immune correlate in vaccinees. In addition, the greater cytotoxic capacity induced in vaccine trials among participants with protective alleles is consistent with associations between MHC alleles and viral load in vaccinees. Over the coming years we are optimistic that this work will provide a long-sought after T cell correlate of immunity that will predict immunologic control of HIV and, in the process, accelerate HIV vaccine development.